Pituitary gland dysfunction affects growth, fertility, the stress response and many other bodily functions. Hypopituitarism can result from congenital defects in organ development (about 1/4000 births) and from pituitary adenomas, which are among the most common type of intracranial tumor. Treatment involves hormone replacement therapy, which can involve daily injections of recombinant growth hormone at great expense per patient. Some adenomas respond to pharmacological therapy, while others are recurrent, potentially causing disfigurement, multiple trans-sphenoidal surgeries, and can result in blindness and death. Genetically engineered mice have proven the roles of several transcription factors and signaling molecules in differentiation, and the correspondence with human pituitary disease is outstanding. In previous years of this grant we defined the roles of four transcription factors in pituitary development and function: PITX2, GATA2, FOXL2, and LHX3. Taken together our analysis of these transcription factors has advanced our understanding of the genetic hierarchy that controls pituitary organogenesis. To understand the mechanisms that underlie congenital pituitary hypoplasia, links need to be made between transcriptional regulators and cell cycle regulators. We established a mouse knockout model of pituitary adenoma that causes recruitment of precursors of the Pou1f1 (PIT1) lineage to produce excess thyrotropes, the pituitary cells that produce thyroid stimulating hormone, at the expense of cells that make growth hormone and prolactin. Little is known about the mechanism of basic thyrotrope specification or the hypertrophy and hyperplasia. The long-term goal of this research is to improve diagnosis and therapy for people with genetic or acquired hypopituitarism by understanding how transcription factors regulate the transition from cell proliferation to differentiation and by understanding the process of stem cell differentiation into hormone producing cells. The overall objective of this application is to determine: (Aim 1) how mutations in the transcription factors Pitx2, Lhx3, Lhx4 and Isl1 cause pituitary hypoplasia by disrupting the cell cycle and altering the transition from precursor cell proliferation to differentiation, (Aim 2) what factors regulate differentiation of the Pou1f1 lineage into thyrotropes, and (3) how selected factors that are enriched in the thyrotrope adenoma transcriptome will exhibit functional enhancement of differentiation in an embryonic stem cell system. The proposed research is innovative because it integrates information gleaned from several different hypoplasia and hyperplasia models and because it will establish a new stem cell differentiation system. This research will be significant because it will link transcription factor defects causing hypoplasia and congenital hypopituitarism to target genes in the cell cycle revealing molecular mechanisms that control the transition from proliferation to differentiation in normal and abnormal development. This information will be informative for identifying the basis for human pituitary diseases including congenital hormone deficiencies and common adenomas.